SUMMARY Antiretroviral therapy (ART) has increased the life expectancy of human subjects infected with the human immunodeficiency virus (HIV), but many now suffer from an increased frequency and earlier onset of vascular events whose mechanisms are not yet clear. We reported reduced endothelium derived relaxation factor (EDRF) and nitric oxide (NO) in isolated living subdermal microvessels from otherwise risk-free young women with well- controlled HIV. The perivascular adipose tissue (PVAT) surrounding blood vessels normally enhances EDRF/NO. However, HIV and ART both can induce lipoatrophy and adipose inflammation and might thereby impair the augmented EDRF/NO response from PVAT. We found that incubation of vascular cells with HIV Tat protein or tenofovir (component of ART) increased cellular senescence signaling and microRNA-34a. How dysfunction of the endothelium, PVAT and miRNAs contribute to premature microvascular aging is unknown, but our results suggest it may entail increased reactive oxygen species (ROS) which we have detected in the blood vessels and PVAT of HIV positive subjects. We will test the hypothesis that HIV infection and/or ART generate ROS to transcribe specific miRNAs that engage the cellular senescence pathway in microvascular endothelium and PVAT. This impairs endothelial function and NO generation and limits their augmentation by adiponectin and other adipokines release from PVAT. We propose to recruit five groups of women from the Washington DC Women's Interagency HIV Study (WIHS) program or Georgetown University Hospital HIV Clinic: HIV negative, young (21-45 years, group 1) and old (50-80 years, group 2) controls and HIV positive women receiving ART, young (21-45 years, group 3) and old (50-80 years, group 4), or young HIV negative women who are receiving pre-exposure prophylaxis (PrEP, group 5; Aim 2 only). Aim 1 (ex vivo, group 1-4) will evaluate the specific pathways and molecules that mediate accelerated age-dependent endothelial dysfunction in the microvessels and PVAT of HIV positive women and will seek the mediators and adipokines responsible. Aim 2 (in vivo, group 1-5) will use non-invasive iontophoretic application of acetylchonline and measurements of cutaneous blood flow using Laser Doppler Flowmetry ( LDF). It will assess endothelial function in dermal blood vessels to evaluate the separate effects of aging, HIV and ART (using HIV negative women receiving ART for PrEP). The two aims will incorporate a novel concept from preliminary cellular and clinical studies that specific microRNAs provide a biological clock of cellular senescence with which to calibrate rapid microvascular aging in HIV. This will be the first study to explore premature aging in microvessels and its external regulation by PVAT and microRNAs in HIV positive individuals and may identify candidate targets for subsequent clinical trials intervention.